This project addresses the use of Hsp90 molecular chaperone -inhibitors to promote cancer cell death. The studies are focused on the mechanisms by which Hsp90 inhibitors synergize when combined with specific protein kinase inhibitors to promote apoptosis. Benzoquinoid ansamycins, including geldanamyin (GA), are compounds that inhibit Hsp90's ATPase activity and promote degradation of client kinases and transcription factors via the ubiquitin/proteasome system. Recent clinical trials with a derivative of GA, 17-AAG, showed that it is well tolerated. Furthermore, 17-AAG appears to be especially effective in promoting death of tumor cells compared with cells from healthy tissue. Hsp90 represents only one of many proteins that have been identified in a new wave of 'targeted'chemotherapeutics. Others include protein kinases (the prototype of targeted therapy), histone deacetyltransferases, the proteasome and anti-apoptotic proteins. In addition, there are many examples where combining therapies towards two targets promotes synergistic cancer cell killing. The mechanisms underlying this synergy likely reflects the way signaling pathways in general are organized into networks, whose robust character can withstand loss of a single component. Since Hsp90 has a general role in buffering signaling pathways, it could provide a basis for chemosensitizing cells to other drugs. Our studies are based on this rationale. In the first aim we will determine the specificity with which casein kinase II (CK2) and Hsp90 inhibitors promote apoptosis in cancer cells, based on preliminary studies. We will characterize the mechanisms of this effect by investigating pathways where CK2 and Hsp90 have a convergent function in cell survival. In aim 2, we will identify additional protein kinases whose loss of function promotes apoptosis in the presence of Hsp90 inhibitors and characterize their roles in cell survival. Finally, we will examine how Akt sensitivity to Hsp90 inhibitors is modulated by cellular environment and how mutation in B-Raf affects its chaperone-dependence.